Restriction and control of HDV infection in stem cell derived hepatocytes.

• Applicant: Dr. Arnaud Carpentier
• Subject Area: Virology
• Term: since 2025
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 539454614

The Hepatitis D Virus (HDV) is a deficient virus necessitating the Hepatitis B Virus (HBV) to complete its viral cycle. HDV-HBV coinfection leads to most severe form of viral hepatitis, with no curative therapy. Understanding the host pathogen interactions of HDV may hold the key to developing new strategies of viral control. In addition to rare primary hepatocytes (PHHs) and poorly relevant transformed hepatoma cell lines (Huh7), we recently described stem cell derived hepatocyte (HLCs) as a new model to study HDV mono-infection. Similarly to PHHs, HLCs support only a low percentage of infected cells. The present project intend to explore the mechanisms of viral restriction and control of HDV in those relevant HLCs. We first intend to complete HDV replication in our HLCs by establishing parameters of HBV and HDV co-infection. We will also use HBV surface antigens (HBsAg) mRNA transfection in our HLCs to complete HDV replication without additional interference from HBV. We will then investigate immune activation of HDV, HBV and co-infected HLCs, and study how the IFN based immune response may control the two viral infections. We will explore the effect of two IFN-independent restriction factors on HDV: CD302 and IRF1. CD302 is a c-type lectin receptor restricting Hepatitis C and E viruses infection in hepatocytes. CD302 is expressed at the same level in HLCs and PHHs, and may thus also control HDV infection of our HLCs. Interferon-regulated factor 1 (IRF1) constitutive expression mediates a basal antiviral state in PHHs. Contrary to deficient Huh7, HLCs express level of IRF1 close to PHHs. Modulating both these factors in highly permissive Huh7 and in relevant HLCs, we intend to assess their antiviral potential on HDV replication. Finally, we will use single cell RNA sequencing to investigate both viral tropism and cellular response to the infection in control HLCs, HDV mono infected and HDV-HBV co-infected HLCs. Detection of HDV and HBV mRNAs will give us the single cell’s viral status. Cellular genes specifically enriched in infected cells will be studied in an attempt to identify new host factors facilitating infection. We will also study the innate immune response of the HLCs at the single cell level. We will visualize immune activation based on replication of one or the other virus. We will compare immune signature of bystander cells vs control non-inoculated cells to assess paracrine IFN stimulation from infected cells.

• DFG Programme: Research Grants